AN IMPROVED SITE-DIRECTED MUTAGENESIS PROCEDURE AND APPLICATION FOR IDENTIFICATION OF MUTANT CLONES

文献类型： 外文期刊

第一作者： Yi, Ganjun

作者： Yi, Ganjun

作者机构：

关键词： OE-PCR;SI nuclease;site-directed mutagenesis

期刊名称：BIOTECHNOLOGY & BIOTECHNOLOGICAL EQUIPMENT （ 影响因子：1.632； 五年影响因子：2.029 ）

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年卷期：

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收录情况： SCI

摘要： A number of molecular tools have been developed to study pathogenicity in fungal phytopathogens, including Fusarium species. Here, we reported an improved method for site-directed mutagenesis to create constructs for using in the analysis of fungal gene function. The procedure combining overlap-extension PCR (OE-PCR) with SI nuclease mismatch cleavage, which we refer to as OE-PCR-S1, is more time- and labor-efficient. We tested our method by deleting three bases from the pathogenic gene Fowl (EU795421) and the resulting mutant DNA was inserted into an expression vector before transformation into Foc for functional domain analysis. The overall rate of mutant site production was 100% and the entire process was completed in less than two days. These results demonstrate that this method is a powerful tool for studying the functions of pathogenic genes and genetic engineering.

分类号： Q

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